The long term goals are:(i) to employ catalytic antibodies to define the fundamental relationships between substrate binding and its catalytic chemical conversion, (ii) to determine whether antibody-mediated catalytic hydrolysis of cell surface antigens, hormones and neurotransmitters is a fundamental pathogenetic mechanism in autoimmune disease, and (ii) to investigate the promise that catalytic anti-peptide antibodies hold as specific anti-viral and anti-tumor therapeutic agents. The specific objectives of the proposed studies are: (i) to demonstrate catalytic hydrolysis of the neurotransmitter VIP by human autoantibodies, (ii) to define the clonotype of the VIP autoantibodies and to purify the VIP- specific antibodies to homogeneity, (iii) to identify the scissile peptide bond(s) and compare the site of hydrolysis with the epitope(s) bound by the antibodies, (iv) to define the sequence specificity of the antibodies based on binding and hydrolysis studies, (v) to measure the frequency of catalytic VIP autoantibodies in defined human subject populations, and to evaluate the bioactivity of fragments produced by antibody-mediated VIP hydrolysis, (vi) to investigate whether the sensitivity of certain peptide bonds to hydrolysis arises from an intrinsic instability of these bonds, (vii) to determine whether known peptidase inhibitors influence antibody- mediated hydrolysis of VIP, (viii) to measure the ability of presumed transition state analogs to bid the antibodies and inhibit hydrolysis of VIP, (ix) to develop human hybridoma cell lines producing monoclonal catalytic anti-VIP antibodies. The experimental design is: (i) purification of mono(125l,Tyr10)-VIP by reverse phase HPLC, (ii) measurement and analysis of mono (125l,Tyr10)-VIP binding by radioimmunoassay and mono(125l,Tyr10)-VIP hydrolysis by trichloracetic acid precipitation and HPLC, (iii) purification of the antibodies by ligand specific affinity chromatography and isoelectric focusing, (iv) covalent crosslinking of antibody:mono(125l,Tyr10) VIP complexes and analysis by two dimensional electrophoresis, (v) localization of antibody binding and hydrolytic activity in gels by novel blotting techniques, (vi) purification of VIP fragments by HPLC and their identification by peptide sequencing and fast atom bombardment-mass spectroscopy, (vii) localization of the antibody binding epitopes by measuring the reactivity of synthetic VIP subsequences in radioimmunoassay, (vii) measurement of antibody specificity using peptides bearing partial sequence identify with VIP in binding and hydrolysis assays, (ix) measurement of binding and catalytic activity of VIP autoantibodies in asthmatics and healthy subjects, (x) investigation of reaction mechanisms by evaluating the instability of peptide bonds, the effect of known inhibitors of the various classes of proteases, on the antibody-mediated hydrolysis, and the reactivity of presumed transition state analogs, by the methods stated above, and (xii)construction of human cell lines making monoclonal catalytic anti-VIP antibody by lymphocyte transformation with Epstein-Barr virus, fusion with myeloma cells, and cloning by limiting dilution techniques.